Abstract Infection with M. tuberculosis (Mtb) can cause TB, which is now the leading global cause of mortality due to a single infectious disease agent. The failure to control TB stems from the lack of an effective vaccine. BCG, the vaccine currently in use, is unable to generate long-lived memory responses and protect against pulmonary TB in adults. New vaccines against TB are therefore, urgently needed. Our work has shown that the ?sigH mutant of Mtb is attenuated for replication and disease in macaques). Aerosol vaccination with this mutant induces strong lung immune signatures that protect against lethal TB. Nonpathogenic ?sigH infection in macaques is not reactivated by SIV co-infection. Additional unrelated mutation(s) are however needed in ?sigH to ensure its complete attenuation. This project aims to develop a ?sigH -based human TB vaccine candidate that could eventually advance to clinical development. ?sigH deletion will be added to 10 recombinant Mtb strains. Some of these mutants generate immune enhancement- or auxotrophy-based attenuation phenotypes, while others render Mtb avirulent in macaque lungs. We will assess the safety of these ten strains in SCID mice and progressively smaller numbers in immunocompetent and SIV co-infected macaques by multiplexing. Two strains that consistently provide the best results in terms of safety/immunogenicity will be evaluated in detail for immunogenicity and efficacy via aerosol (AER) vaccination relative to BCG- and ?sigH - vaccination, in a physiologically relevant macaque model.